Proper nervous system function depends on precise contacts among neurons, and between neurons and their non-neuronal targets. My goal is to learn how neurons find and recognize their targets during development. I plan to examine identified motoneurons in live zebrafish embryos to learn how interactions among their growth cones and between their growth cones and other cells influence the cell-specific pathways they select. During development, the growth cones of the three identified motoneurons in each spinal segment of the zebrafish leave the spinal cord in an invariant sequence in which the growth cone of the first motoneuron acts as a pioneer in establishing the pathway followed by later motor growth cones. I will test whether the other motoneurons can function as pioneers by ablating the normal pioneer motoneuron prior to axogenesis with a dye-pumped pulse laser. I will look for interactions among identified motor growth cones that may function in pathway and target selection. First, I will determine whether identified motor growth cones can follow alternate pathways by ablating identified motoneurons and observing the behavior of the remaining motoneurons. Second, I will delay the growth cone of an identified motor neuron by ablating it without killing the rest of the cell, and I will observe whether this alters growth cone pathway choice. My results will help to determine whether growth cones compete for pathway or targets. I will examine whether coupling among identified motoneurons is important in pathway selection by labeling them with low molecular weight fluorescent dyes and observing the time-dependence of coupling and uncoupling. I will also prevent normal, time- and activity-dependent uncoupling by manipulating neuronal activity, to determine whether uncoupling is required for pathway choice. I will investigate interactions between identified motor growth cones and their pathways using electron microscopy to examine particular pathway features that may serve as guidance cues.